Source attribution of salmonellosis by time and geography in New South Wales, Australia.

BACKGROUND: Salmonella is a major cause of zoonotic illness around the world, arising from direct or indirect contact with a range of animal reservoirs. In the Australian state of New South Wales (NSW), salmonellosis is believed to be primarily foodborne, but the relative contribution of animal reservoirs is unknown. METHODS: The analysis included 4543 serotyped isolates from animal reservoirs and 30,073 serotyped isolates from domestically acquired human cases in NSW between January 2008 and August 2019. We used a Bayesian source attribution methodology to estimate the proportion of foodborne Salmonella infections attributable to broiler chickens, layer chickens, ruminants, pigs, and an unknown or unsampled source. Additional analyses included covariates for four time periods and five levels of rurality. RESULTS: A single serotype, S. Typhimurium, accounted for 65-75% of included cases during 2008-2014 but < 50% during 2017-2019. Attribution to layer chickens was highest during 2008-2010 (48.7%, 95% CrI 24.2-70.3%) but halved by 2017-2019 (23.1%, 95% CrI 5.7-38.9%) and was lower in the rural and remote populations than in the majority urban population. The proportion of cases attributed to the unsampled source was 11.3% (95% CrI 1.2%-22.1%) overall, but higher in rural and remote populations. The proportion of cases attributed to pork increased from approximately 20% in 2009-2016 to approximately 40% in 2017-2019, coinciding with a rise in cases due to Salmonella ser. 4,5,12:i:-. CONCLUSION: Layer chickens were likely the primary reservoir of domestically acquired Salmonella infections in NSW circa 2010, but attribution to the source declined contemporaneously with increased vaccination of layer flocks and tighter food safety regulations for the handling of eggs.

Shiga toxin-producing Escherichia coli in ground beef and lamb cuts: results of a one-year study.

Shiga toxin-producing Escherichia coli (STEC) have been associated with a broad spectrum of diarrhoeal syndromes. Some of these cases have been attributed to foods of bovine origin or other foods cross-contaminated by beef products or cow manure. The purpose of this study was to determine the pattern of STEC distribution in selected red meats over time. Samples of ground beef and lamb cuts were collected over a 52-week period from 31 different outlets and 25 g portions were assayed for STEC. STEC were isolated from 46/285 (16%) ground beef and 111/275 (40%) lamb samples using an stx PCR screen followed by colony hybridisation. All isolates were tested by PCR for additional STEC virulence markers with 95% of ground beef isolates shown to possess stx(2) and 80% of lamb cutlet isolates shown to possess stx(1) and stx(2). The enterohaemolysin gene (ehxA) was detected in 65% and 53% of ground beef and lamb isolates respectively. Putative enterohaemorrhagic E. coli (EHEC), i.e. STEC possessing the E. coli attaching and effacing gene (eae) were not isolated. The STEC isolates comprised 18 and 15 different serotypes from ground beef and lamb respectively. STEC of serotypes O157, O111 and O26 (common enterohaemorrhagic E. coli serotypes) were not isolated. Serotypes O174 and O91 were the most common serotypes isolated from ground beef samples and O128 and O91 the most common from lamb cutlet samples. The presence of STEC in retail red meats highlights the need for a clearer understanding of STEC in food and human illness to interpret the public health significance of these findings.

Isolation and characterization of integron-containing bacteria without antibiotic selection.

The emergence of antibiotic resistance among pathogenic and commensal bacteria has become a serious problem worldwide. The use and overuse of antibiotics in a number of settings are contributing to the development of antibiotic-resistant microorganisms. The class 1 and 2 integrase genes (intI1 and intI2, respectively) were identified in mixed bacterial cultures enriched from bovine feces by growth in buffered peptone water (BPW) followed by integrase-specific PCR. Integrase-positive bacterial colonies from the enrichment cultures were then isolated by using hydrophobic grid membrane filters and integrase-specific gene probes. Bacterial clones isolated by this technique were then confirmed to carry integrons by further testing by PCR and DNA sequencing. Integron-associated antibiotic resistance genes were detected in bacteria such as Escherichia coli, Aeromonas spp., Proteus spp., Morganella morganii, Shewanella spp., and urea-positive Providencia stuartii isolates from bovine fecal samples without the use of selective enrichment media containing antibiotics. Streptomycin and trimethoprim resistance were commonly associated with integrons. The advantages conferred by this methodology are that a wide variety of integron-containing bacteria may be simultaneously cultured in BPW enrichments and culture biases due to antibiotic selection can be avoided. Rapid and efficient identification, isolation, and characterization of antibiotic resistance-associated integrons are possible by this protocol. These methods will facilitate greater understanding of the factors that contribute to the presence and transfer of integron-associated antibiotic resistance genes in bacterial isolates from red meat production animals.

Presence of activatable Shiga toxin genotype (stx(2d)) in Shiga toxigenic Escherichia coli from livestock sources.

Stx2d is a recently described Shiga toxin whose cytotoxicity is activated 10- to 1000-fold by the elastase present in mouse or human intestinal mucus. We examined Shiga toxigenic Escherichia coli (STEC) strains isolated from food and livestock sources for the presence of activatable stx(2d). The stx(2) operons of STEC were first analyzed by PCR-restriction fragment length polymorphism (RFLP) analysis and categorized as stx(2), stx(2c vha), stx(2c vhb), or stx(2d EH250). Subsequently, the stx(2c vha) and stx(2c vhb) operons were screened for the absence of a PstI site in the stx(2A) subunit gene, a restriction site polymorphism which is a predictive indicator for the stx(2d) (activatable) genotype. Twelve STEC isolates carrying putative stx(2d) operons were identified, and nucleotide sequencing was used to confirm the identification of these operons as stx(2d). The complete nucleotide sequences of seven representative stx(2d) operons were determined. Shiga toxin expression in stx(2d) isolates was confirmed by immunoblotting. stx(2d) isolates were induced for the production of bacteriophages carrying stx. Two isolates were able to produce bacteriophages phi1662a and phi1720a carrying the stx(2d) operons. RFLP analysis of bacteriophage genomic DNA revealed that phi1662a and phi1720a were highly related to each other; however, the DNA sequences of these two stx(2d) operons were distinct. The STEC strains carrying these operons were isolated from retail ground beef. Surveillance for STEC strains expressing activatable Stx2d Shiga toxin among clinical cases may indicate the significance of this toxin subtype to human health.

Enterohemorrhagic Escherichia coli -The Australian Perspective †.

Food borne transmission of hemolytic uremic syndrome (HUS) was first reported in Australia in 1995 when an outbreak of HUS due to Escherichia coli O111 occurred following the consumption of locally produced mettwurst. Federal and state health and food authorities responded rapidly to bring the outbreak under control. Longer-term responses include the introduction by regulatory authorities of a code of practice for uncooked fermented comminuted meat products, the provision of government and industry funds to support the implementation of this code, and research into the ecology and epidemiology of enterohemorrhagic Escherichia coli and the safe production of meat. In addition, general awareness has increased, and activities in food safety control among all sectors has been stimulated. The pattern of EHEC serotypes in the Australian human and animal populations appears different from that in countries in the Northern Hemisphere. Serotype O157:H7 is not the predominant serotype isolated. Other serotypes, including O111, are more common and possess a variety of virulence-associated determinants. Research into food safety and EHEC is therefore aimed at the development of detection methods more appropriate for the Australian situation. Additional research objectives include determining both the prevalence of EHEC in meat and the meat animal population and farming and handling practices that influence EHEC carriage and transmission. These activities will contribute to an assessment of the hazards presented by EHEC in Australia and recommendations for their control.